1. Use of Gen-Probe APTIMA Tests to Detect Multiple Etiologies of
Urethritis and Cervicitis
In Sexually Transmitted Disease Clinics
C-086
Board 133
Charlotte A. Gaydos, Andrew Hardick, Justin Hardick, Mathilda Barnes, Billie Jo Wood
Johns Hopkins University, Baltimore, MD

2. Abstract
Background: In addition to Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (GC), being commonly recognized causes of urethritis and cervicitis, other agents, such as Trichomonas vaginalis (TV) and Mycoplasma genitalium (MG), can cause urethritis and cervicitis. The Gen-Probe APTIMA TMA technology can be used to detect each of the 4 pathogens. Methods: Urines from 290 men and self-administered vaginal swabs from 325 women were collected from STD clinic patients. Demographics, behavioral risk factors, signs, and symptoms were recorded. CT and GC were detected by APTIMA Combo 2. MG and TV were detected by real time research PCR-MG and B-TUB FRET-TV and by prototype Gen-Probe TMA-based assays for MG and TV (TMA-MG and TMA-TV). A patient was considered to be infected with MG and/or TV if both research PCR assay and prototype TMA assay were positive. Results: Prevalence: males (N=153) with urethritis: CT 32.7%, GC 24.2%, TV 5.2%, MG 22.4%; males (N=137) without urethritis: CT 6.6%, GC 0%, TV 1.5%, MG 7.3%. Prevalence: females (N=133) with cervicitis: CT 15.4%, GC 6.0%, TV 18.9%, MG 28.6%; females (N=192) without cervicitis: CT 8.3%, GC 3.7%, TV 12.7%, MG 13.2%. Kappa statistic comparison of PCR and TMA-MG and TMA-TV were excellent (0.941 and 0.858, respectively). Of infected women and men, 23.6% and 20.3% had >1 pathogen, respectively. In regression analysis, women with symptoms were more likely to have cervicitis (p=0.04) and men who were contacts of persons with STDs were more likely to have urethritis (p=0.05). Only MG was associated with cervicitis and all organisms except TV were associated with urethritis in men. Conclusion: Prevalence of any organism was higher in urethritis and cervicitis. Co-infections were common. TMA-based assays for MG and TV performed very well compared to research PCRs.

3. Introduction Definition: Cervicitis: Inflammation of the cervix
Urethritis: Inflammation of the urethra
Diagnosis: can vary
Cervicitis:
Discharge, friability, gram stain PMNs
Mucopurulent endocervical discharge (yellow exudate)
Appearance of the inflamed endocervix on physical exam (discharge, edema, erythema)
Urethritis:
Urethral GS > 5 PMNs/HPF
Urine sediment GS: 15/400X; Leukocyte Esterase

4. Urethritis and Cervicitis
Incidence:
Urethritis (CDC) over 4 million cases per year
Gonococcal- 650,000 cases/yr
Non Gonooccal-over 2 million cases/ yr
Up to 65% are of unknown origin
Cervicitis- estimated 4 million cases per year
Reported to CDC, CT/GC 832,506 cases
Up to 40% are of unknown etiology

5. Signs, Symptoms, Risk Factors
Males
Urethral discharge
Dysuria
Itching at end of urethra
Females
Discharge
Abnormal vaginal bleeding
Abdominal pain
Mucopus
Young age
Multiple/new partners
Sex/social networks
Hx STD
Race
Geography
Age 1st sex

6. Objectives
To determine the prevalence and multiple etiologies of male urethritis and female cervicitis
To determine the prevalence of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (GC), Mycoplasma genitalium (MG) and Trichomonas vaginalis (TV) in patients from STD clinics
To compare the research Gen-Probe APTIMA TMA for MG and TV with research PCRs.
To determine the association of CT, GC, MG, and TV with cervicitis and urethritis

7. Mycoplasma genitalium
Thought to be sexually transmitted; also found in respiratory tract
Associated with NCNGGU in men and cervicitis in women recently
Possible sequelae
endometritis?, PID?
adverse birth outcomes??
Smallest prokaryote, with no cell wall, and very difficult to culture

8. Trichomonas vaginalis
More prevalent than CT or GC: CDC estimates 5 million cases/yr in U.S.; Not a reportable disease
Men: may be asymptomatic (>50%),
NCNGGU, may cause prostatitis,
epididymitis; assoc w/ significant
decrease in sperm motility and viability,
isolated from 10% infertile men
Women: asymptomatic (<50%) or cause frothy discharge, risk for cervical neoplasia, tubal infertility, post-hysterectomy infection, atypical PID preterm birth, low birth weight
Risk factor for HIV transmission

9. Methods Men (290) and women (325) attending STD clinics
Urethritis in men was dx’d by >5 PMN on Gram Stain
Cervicitis in women was dx’d by cervical mucopus, friability, or discharge
Men provided 2 urethral swabs, urine;
Women provided 3 cx swabs, 2 vaginal swabs, urine
Routine tests included APTIMA Combo2 (GenProbe) tests for CT and GC from male urine, female cervical, vaginal and urine samples

10. Methods
Research PCRs were performed for T. vaginalis and M. genitalium on male urine and female vaginal samples
Research APTIMA TMA- based assays (GenProbe) were performed for M. genitalium (TMA-MG) and for T. vaginalis (TMA-TV) on male urine and female vaginal samples
A patient was considered to be infected with MG and/or TV if both research PCR assay and research TMA assay were positive.

11. Methods Trichomonas Research PCR*
based on detection of the B tubulin gene
real-time PCR Roche Light Cycler technology
Mycoplasma Research PCR**
utilized 2 targets in a diplex assay
MgPa adhesion gene
16S rRNA gene
7900 Sequence Detection ABI Prisim
*Hardick et al. JCM 41: , 2003; *Madico et el. JCM 36: , 1998
** Hardick et al. JCM 44: , 2006

12. Microbial Etiology Study Results: N= 325 females, 290 malesCT
GC 4.6
TV 15.3
MG None 60.9
Males: %
CT 20.3
GC 12.8
TV 3.4
MG 15.2
None 57.6
Median Age 24 yr.
Black 92.6%
Symptoms 69.8%
Median Age 25 yr
Black 96.9%
Symptoms 66.9%

13. Results Females: Males Urethritis N=153 No urethritis N=137
Cervicitis N=133 No cervicitis N=192
CT 15.0% %
GC 6.0% %
TV 18.8% %
MG 28.6% %
Males
Urethritis N=153 No urethritis N=137
CT 32.7% %
GC 24.2% %
TV 5.2% %
MG 22.4% %

14. Results
Kappa statistics for comparison between research PCRs for MG and TV and TMA-MG and TMA-TV were excellent (0.941 and 0.858, respectively)
Co-infections of > 2 organisms occurred in 10.6% of all males; if infected with at least one pathogen, % had a coinfection
Co-infections with > 2 organisms occurred in 11.4% of all females; if infected with at least one pathogen, 30% had a coinfection

15. Association with Cervicitis
Univariate Multivar Model 1 Model 2
OR p value OR p OR p CT NG TV MG
Age <25 yr
Contact

16. Association with Urethritis
Univariate Multivariate Model 1
OR p value OR p
CT 6.9 < <0.0001
NG* < * TV MG
Age <25 yr
Contact
*NG could not be fit into the multivariate model due to collinearity between NG and urethritis that caused the model not to fit

17. Urethritis Infection in men with urethritis
Infection in men without urethritis
33.1%

18. Cervicitis Coinfection in women with cervicitis
Coinfection in women without cervicitis
47.4%
72.3%

19. Conclusions
Prevalence of all organisms was higher in men with urethritis and in women with cervicitis in this population
Co-infections were common; MG and TV were associated with urethritis and cervicitis
Research TMA-based GenProbe assays for MG and TV performed very well compared to research PCR assays
Use of newer tests such as TMA or PCR will allow better etiologic diagnostic capability in STD patients
Even with increased diagnostic capability, a significant proportion of urethritis and cervicitis cases have no etiologic agent detected